The membrane bound electron transport complex of Micrococcus lysodeikticus has been disrupted into a soluble fraction containing NADH and L-malate dehydrogenase enzymes and a particulate vesicular fraction containing cytochromes a, b, and c. Reconstitution of NADH and L-malate oxidases has as its basis a physical reconstitution of solubilized dehydrogenases with particulate dehydrogenase depleted membranes. The proposed work will establish whether any solubilized coupling factors are required for reconstitution of purified dehydrogenase enzymes. Particulate, dehydrogenase depleted, membrane vesicles will be solubilized and components separated to determine which components are required for reconstituting the complete multi-enzyme oxidase complex. These methods will be applied to studying differentiation of Staphylococcus membranes during conversion of growing cells from a fermentative to a respiratory metabolism. Transport of glucose in Pseudomonas fluorescens has been shown to involve uptake of glucose carbon against a concentration gradient (active transport). A glucose oxidase deficient mutant of this organism cannot accumulate glucose carbon via active transport. Studies proposed will establish the specific mechanism(s) of sugar transport in this bacterium which appears to have a novel substrate dependent membrane oxidase for substrate (glucose) transport. The roles of membrane oxidases in this organism will be further studied to establish how these membrane oxidases regulate cellular activities.